单纯疱疹病毒Ⅰ型糖蛋白B胞浆区编码基因的原核表达、纯化及抗体制备  

Cloning and prokaryotic expression, purification and amplification to preparing specific antibody of gene encoding herpes simplex virus typeⅠglycoprotein B cytoplasmic domain

作　　者：李英辉[1] 刘军[1] 薛采芳[1] 甄荣芬[1] 李旬[1] 王宪锋[1] 刘忠湘[1] 万磊[1] 

机构地区：[1]第四军医大学病原生物学教研室,陕西西安710032

出　　处：《细胞与分子免疫学杂志》2001年第6期549-551,共3页Chinese Journal of Cellular and Molecular Immunology

基　　金：国家自然科学基金资助;No.39970038

摘　　要：目的获得高纯度的重组单纯疱疹病毒Ⅰ型HSV-1糖蛋白BgB胞浆区蛋白并制备其特异性抗体。方法从感染HSV-1的BHK细胞中提取总RNA用RT-PCR特异性扩增HSV-1gB胞浆区编码基因,经双酶切后,克隆入表达载体pGEX4T-1中,进行融合表达。以纯化的重组蛋白免疫小鼠制备抗体,进行Westernblot鉴定。结果用IPTG诱导后,表达出相对分子质量Mr约42000的融合蛋白。用纯化的融合蛋白免疫小鼠制备的抗体滴度为1400。结论获得重组HSV-1gB胞浆区蛋白及其特异性抗体,为后续功能研究奠定了基础。Aim To obtain recombinant herpes simplex virus type Ⅰ(HSV-1) glycoprotein B(gB)cytoplasmic domain protein with high purity and to prepare specific antibody. Methods The total RNA were extracted from BHK cells infected with HSV-1 virus(SM44) and the gene encoding HSV-1 gB cytoplasmic domain amplified by RT-PCR was cloned into expression vector pGEX4T-1. The recombinant plasmid was expressed in E.coli and the purified recombinant protein was used to immunize the Balb/c mice to prepare specific antibody. Obtained antibody was analyzed by Western blot. Results After IPTG induction, relative molecule mass(Mr) of expressed fusion protein was 42 000. The antitody titer from the Balb/c mice immunized by purified product was 1:400. Conclusion The recombinant HSV-1gB cytoplasmic domain protein is obtained and specific antibody is prepared successfully, thus lay the foundation for further research.

关 键 词：单纯疱疹病毒I型 糖蛋白B 原核表达 谷胱甘肽巯基-转移酶 纯化 抗体 

分 类 号：R392－33[医药卫生—免疫学] R373.11[医药卫生—基础医学]